Spinal muscular atrophy (SMA) is a neurological disorder that results from loss of function of the anterior horn cells in the spinal cord, manifesting as progressive motor weakness, muscle wasting, and paralysis. SMA is caused by insufficient levels of the survival motor neuron (SMN) protein. The SMN locus on chromosome 5q13 contains two inverted copies of SMN called SMN1 and SMN2. Most cases of SMA harbor homozygous deletions of the SMN1 gene and retain at least one copy of SMN2. With a carrier rate of about 1 in 40, SMA is estimated to be the most frequent genetic cause of infant mortality.
SMN2 is a gene duplication of SMN1 with the same predicted amino acid coding capacity. The nucleotide sequences of SMN1 and SMN2 are nearly identical. A critical difference is a C to T transition at the +6 position in exon 7, which dramatically influences the splicing pattern in these genes. Greater than 90% of SMN1 transcripts include exon 7, while there is less than 15% exon 7 inclusion in SMN2 transcripts. This alternatively spliced product produces a truncated and unstable form of the SMN protein. Any increase in the inclusion of exon 7 in SMN2 transcripts would result in higher levels of full length SMN protein, particularly, just doubling the amount of full length SMN2 mRNA could be clinically significant. A treatment that increases the amount of full length SMN2 mRNA should result in increased levels of SMN protein. It is believed that candidate drugs for treating SMA should (i) significantly increase cellular levels of SMN protein expression from the SMN2 gene; (ii) give consistent plasma and brain exposure in mouse (and predicted for human); and (iii) be efficacious in SMA mouse models. Based on this premise, an in vivo screen that can detect increases in full-length exon 7 included SMN2 transcripts was developed.
Current therapeutic strategies for SMA are mostly centered on elevating full length (wild type) SMN protein levels, modulating splicing towards exon 7 inclusion, stabilizing the wild type protein, and to a lesser extent, on restoring muscle function in SMA by providing trophic support or by inhibiting skeletal muscle atrophy. The mechanism leading to motorneuron loss and to muscular atrophy still remains obscure, although the availability of animal models of the disease is rapidly increasing knowledge in this field (Frugier T, et al. (2000) Hum Mol. Genet. 9:849-58; Monani U R, et al. (2000) Hum Mol Genet 9:333-9; Hsieh-Li H M, et al. (2000) Nat Genet 24:66-70; Jablonka S, et al. (2000) Hum Mol. Genet. 9:341-6). Also the function of SMN protein is still partially unknown, and studies indicate that it can be involved in mRNA metabolism (Meister G, et al. (2002). Trends Cell Biol. 12:472-8; Pellizzoni L, et al. (2002). Science. 298: 1775-9), and probably in transport of proteins/mRNA to neuromuscular junctions (Ci-fuentes-Diaz C, et al. (2002) Hum Mol. Genet. 11: 1439-47; Chan Y B, et al. (2003) Hum Mol. Genet. 12:1367-76; McWhorter M L, et al. (2003) J. Cell Biol. 162:919-31; Rossoll W, et al. (2003) J. Cell Biol. 163:801-812).
Accordingly, there is a need for new drugs to treat spinal muscular atrophy. SMN reporters can be used as tools for identifying and characterizing protein factors and chemical compounds that increase levels of full-length SMN protein through mechanisms, including, for example, increased transcription of SMN2, increased inclusion of exon 7 in the SMN2 mRNA, increased stability of the SMN2 mRNA, and decreased degradation of the full-length SMN protein. Results from high throughput systems to identify compounds that increase SMN protein using this cell based SMN-luciferase reporter assay are described herein. As such, the present disclosure provides compounds, as well as combinations of these compounds, useful for treating or lessening the severity of spinal muscular atrophy. The present disclosure also provides methods of treating or lessening the severity of spinal muscular atrophy comprising administering to a patient susceptible to or having spinal muscular atrophy a compound or composition of the present disclosure.